The most crucial point we will investigate is whether the fraction of subfragment-one (S-1) which neither binds ATP irreversibly nor shows the initial Pi burst is, in fact, denatured. It is of importance to make certain that S-1 preparations can be prepared which have very little of this fraction. Purification of the S-1 or a DEAE cellulose column as well as preparation of S-1 with chymotrypsin rather than papain, should help resolve this question. In addition, heavy meromyosin and myosin itself will be studied. It is also important to make certain that the fraction of S-1 which does not show the initial Pi burst can still bind ATP reversibly. If it does so, it will be of interest to study the ATPase properties of this fraction both with and without actin present. Even if it is denatured, its properties may aid us in understanding the role of initial Pi burst in undernatured myosin. Another aspect of this project which requires further study is our finding that H ion release accompanies the hydrolysis of ATP in the initial Pi burst. It has been reported that the binding of ADP on the ATP analogue AmPPNP also causes release of H ion, and whether this is, in fact, the case will require further investigation.